Temperature resistant mutants of Rhodobacter capsulatus generated by a directed evolution approach and effects of temperature resistance on hydrogen production

Hydrogen (H₂) is a promising alternative energy carrier which can be produced biologically. Rhodobacter capsulatus, a non-sulfur purple photosynthetic bacterium, can produce H₂ under nitrogen-limited, photoheterotrophic conditions by using reduced carbon sources such as simple organic acids. Outdoor closed photobioreactors; used for biological H₂ production are located under direct sunlight, as a result; bioreactors are exposed to temperature fluctuations during day time. In this study to overcome this problem, temperature-resistant mutants (up to 42 °C) of R. capsulatus were generated in this study by a directed evolution approach. Eleven mutant strains of R. capsulatus DSM 1710 were obtained by initial ethyl methane sulfonate (EMS) mutagenesis of the wild-type strain, followed by batch selection at gradually increasing temperatures up to 42 °C under respiratory conditions. The genetic stability of the mutants was tested and eight were genetically stable. Moreover, H₂ production of mutant strains was analyzed; five mutants produced higher amounts of H₂ when compared to the DSM 1710 wild-type strain and three mutants produced less H₂ by volume. The highest H₂- producing mutant (B41) produced 24% more H₂ compared to wild type, and the mutant with lowest H₂-production capacity (A52) generated 7% less H₂ compared to the wild type. These results indicated that heat resistance of R. capsulatus can be improved by directed evolution, which is a useful tool to improve industrially important microbial properties. To understand molecular changes that confer high temperature-resistance and high hydrogen production capacity to these mutants, detailed transcriptomic and proteomic analyses would be necessary.